The purpose of this project is to study immune responses in man and animals to natural and experimental rickettsial infections, and to isolate and identify the causative pathogens. For serological investigations, recently developed procedures such as indirect imunofluorescence, microagglutination, and the enzyme-linked immunosorbent assays (ELISA) are being used. It also provides serologic support to other RML units and occasionally also to outside agencies, and includes serodiagnosis of other bacterial or viral diseases under investigation. For the isolation of pathogens, susceptible laboratory animals (meadow voles, guinea pigs, embryonated hen eggs, etc.) and various tissue culture systems (Vero, L cells, etc.) are being used. Serologic tests as well as immunochemical procedures (SDS-PAGE, western blotting) are applied to identification of isolates. Investigations into a disease of suspected rickettsial etiology that killed two persons and hospitalized six others in Virginia, revealed one person to be infected with a typhus agent. Attempts to isolate the organism from brain tissues, blood clots and spinal fluids of several other patients failed. A bacterial agent of hitherto unknown identity was recovered from the brain tissue of a deceased patient. Diagnostic reagents and serologic confirmation was provided in conjunction with a Q fever outbreak in Bagnes, Switzerland here 191 acute cases of this disease had occurred. Of 2,962 patients without apparent illness, 224 (8%) had antibodies to Coxiella burnetii. A comparative study on the sensitivity between ELISA and indirect immunofluorescent antibody tests in the serologic diagnosis of primary, granulomatous, and endocarditis infections of Q fever in man, revealed good correlation. Q fever endocarditis could regularly be identified by high titers of antibodies to phase I particulate antigen. Nonspecific FA staining reactions in sections of human heart valves infected with C. burnetii was found to be eliminated through pepsin and trypsin enzyme treatment of sections prior to FA staining.